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Occurrence of grapevine leafroll-associated virus complex in Napa Valley.

Sharma AM, Wang J, Duffy S, Zhang S, Wong MK, Rashed A, Cooper ML, Daane KM, Almeida RP - PLoS ONE (2011)

Bottom Line: Furthermore, two novel variants, which grouped with a divergent isolate from New Zealand (NZ-1), were identified, and these variants comprised 6% of all positive GLRaV-3 samples.Spatial analyses showed that GLRaV-3a, 3b, and 3c were not homogeneously distributed across Napa Valley.Overall, 86% of all blocks (n = 31) were positive for GLRaVs and 90% of positive blocks (n = 28) had two or more GLRaV-3 variants, suggesting complex disease dynamics that might include multiple insect-mediated introduction events.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Science, Policy and Management, University of California, Berkeley, California, United States of America.

ABSTRACT
Grapevine leafroll disease (GLD) is caused by a complex of several virus species (grapevine leafroll-associated viruses, GLRaV) in the family Closteroviridae. Because of its increasing importance, it is critical to determine which species of GLRaV is predominant in each region where this disease is occurring. A structured sampling design, utilizing a combination of RT-PCR based testing and sequencing methods, was used to survey GLRaVs in Napa Valley (California, USA) vineyards (n = 36). Of the 216 samples tested for GLRaV-1, -2, -3, -4, -5, and -9, 62% (n = 134) were GLRaV positive. Of the positives, 81% (n = 109) were single infections with GLRaV-3, followed by GLRaV-2 (4%, n = 5), while the remaining samples (15%, n = 20) were mixed infections of GLRaV-3 with GLRaV-1, 2, 4, or 9. Additionally, 468 samples were tested for genetic variants of GLRaV-3, and of the 65% (n = 306) of samples positive for GLRaV-3, 22% were infected with multiple GLRaV-3 variants. Phylogenetic analysis utilizing sequence data from the single infection GLRaV-3 samples produced seven well-supported GLRaV-3 variants, of which three represented 71% of all GLRaV-3 positive samples in Napa Valley. Furthermore, two novel variants, which grouped with a divergent isolate from New Zealand (NZ-1), were identified, and these variants comprised 6% of all positive GLRaV-3 samples. Spatial analyses showed that GLRaV-3a, 3b, and 3c were not homogeneously distributed across Napa Valley. Overall, 86% of all blocks (n = 31) were positive for GLRaVs and 90% of positive blocks (n = 28) had two or more GLRaV-3 variants, suggesting complex disease dynamics that might include multiple insect-mediated introduction events.

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Schematic illustration of sampling design used in this study.Samples were first screened for different GLRaV species, then two approaches were used to type GLRaV-3. Frag. An. stands for ‘Fragment analysis’, a typing approach described in Materials and Methods.
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pone-0026227-g001: Schematic illustration of sampling design used in this study.Samples were first screened for different GLRaV species, then two approaches were used to type GLRaV-3. Frag. An. stands for ‘Fragment analysis’, a typing approach described in Materials and Methods.

Mentions: Based on the results of the species level identification scheme, petioles were further tested to identify specific GLRaV-3 variants. If a block was positive for GLRaV-3, then the previous samples plus additional 5 to 10 samples were tested (Figure 1). However, if a block was negative for GLRaV-3, the five samples tested above were tested again for the presence of GLRaV-3 variants in case the primers designed for this study (described below) detected potential positives that were missed by the primer used for the species level survey. Fluorescently labeled primers (Table S3) were designed to distinguish between the four GLRaV-3 variants identified in Napa Valley, CA by Wang et al. [20]. The same terminology for genetic variants was used in this study. Forward and reverse primers were designed based on multiple alignments that identified regions conserved in one of the four variants but with low sequence similarity in comparison to the other three variants. In order to detect isolates the variant specific primers might miss, a general GLRaV-3 primer set was designed from regions of the coat protein (CP) gene conserved within the 50 isolates in Wang et al. [20] and other sequences deposited in GenBank. This primer set is hereafter designated as CP primer set (Table S3). Primer sets were first tested and assayed individually and then in a multiplex setup using the RNA extracted from a previous project [20].


Occurrence of grapevine leafroll-associated virus complex in Napa Valley.

Sharma AM, Wang J, Duffy S, Zhang S, Wong MK, Rashed A, Cooper ML, Daane KM, Almeida RP - PLoS ONE (2011)

Schematic illustration of sampling design used in this study.Samples were first screened for different GLRaV species, then two approaches were used to type GLRaV-3. Frag. An. stands for ‘Fragment analysis’, a typing approach described in Materials and Methods.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3198396&req=5

pone-0026227-g001: Schematic illustration of sampling design used in this study.Samples were first screened for different GLRaV species, then two approaches were used to type GLRaV-3. Frag. An. stands for ‘Fragment analysis’, a typing approach described in Materials and Methods.
Mentions: Based on the results of the species level identification scheme, petioles were further tested to identify specific GLRaV-3 variants. If a block was positive for GLRaV-3, then the previous samples plus additional 5 to 10 samples were tested (Figure 1). However, if a block was negative for GLRaV-3, the five samples tested above were tested again for the presence of GLRaV-3 variants in case the primers designed for this study (described below) detected potential positives that were missed by the primer used for the species level survey. Fluorescently labeled primers (Table S3) were designed to distinguish between the four GLRaV-3 variants identified in Napa Valley, CA by Wang et al. [20]. The same terminology for genetic variants was used in this study. Forward and reverse primers were designed based on multiple alignments that identified regions conserved in one of the four variants but with low sequence similarity in comparison to the other three variants. In order to detect isolates the variant specific primers might miss, a general GLRaV-3 primer set was designed from regions of the coat protein (CP) gene conserved within the 50 isolates in Wang et al. [20] and other sequences deposited in GenBank. This primer set is hereafter designated as CP primer set (Table S3). Primer sets were first tested and assayed individually and then in a multiplex setup using the RNA extracted from a previous project [20].

Bottom Line: Furthermore, two novel variants, which grouped with a divergent isolate from New Zealand (NZ-1), were identified, and these variants comprised 6% of all positive GLRaV-3 samples.Spatial analyses showed that GLRaV-3a, 3b, and 3c were not homogeneously distributed across Napa Valley.Overall, 86% of all blocks (n = 31) were positive for GLRaVs and 90% of positive blocks (n = 28) had two or more GLRaV-3 variants, suggesting complex disease dynamics that might include multiple insect-mediated introduction events.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Science, Policy and Management, University of California, Berkeley, California, United States of America.

ABSTRACT
Grapevine leafroll disease (GLD) is caused by a complex of several virus species (grapevine leafroll-associated viruses, GLRaV) in the family Closteroviridae. Because of its increasing importance, it is critical to determine which species of GLRaV is predominant in each region where this disease is occurring. A structured sampling design, utilizing a combination of RT-PCR based testing and sequencing methods, was used to survey GLRaVs in Napa Valley (California, USA) vineyards (n = 36). Of the 216 samples tested for GLRaV-1, -2, -3, -4, -5, and -9, 62% (n = 134) were GLRaV positive. Of the positives, 81% (n = 109) were single infections with GLRaV-3, followed by GLRaV-2 (4%, n = 5), while the remaining samples (15%, n = 20) were mixed infections of GLRaV-3 with GLRaV-1, 2, 4, or 9. Additionally, 468 samples were tested for genetic variants of GLRaV-3, and of the 65% (n = 306) of samples positive for GLRaV-3, 22% were infected with multiple GLRaV-3 variants. Phylogenetic analysis utilizing sequence data from the single infection GLRaV-3 samples produced seven well-supported GLRaV-3 variants, of which three represented 71% of all GLRaV-3 positive samples in Napa Valley. Furthermore, two novel variants, which grouped with a divergent isolate from New Zealand (NZ-1), were identified, and these variants comprised 6% of all positive GLRaV-3 samples. Spatial analyses showed that GLRaV-3a, 3b, and 3c were not homogeneously distributed across Napa Valley. Overall, 86% of all blocks (n = 31) were positive for GLRaVs and 90% of positive blocks (n = 28) had two or more GLRaV-3 variants, suggesting complex disease dynamics that might include multiple insect-mediated introduction events.

Show MeSH
Related in: MedlinePlus